人干扰素α2b ELISA配体结合分析检测方法的建立  

Establishment of ELISA based on ligand binding for detection of human interferon alpha 2b

作　　者：代晓阳 张燕[1] 沈丽红 解翠薇 杨晓春[1] 翁勤洁(指导)[1] DAI Xiaoyang;ZHANG Yan;SHEN Lihong;XIE Cuiwei;YANG Xiaochun;WENG Qinjie(Center for Drug Safety Evaluation and Research of Zhejiang University,Hangzhou 310000,China)

机构地区：[1]浙江大学药物安全评价研究中心,杭州310000

出　　处：《中国免疫学杂志》2023年第6期1312-1317,共6页Chinese Journal of Immunology

基　　金：国家自然科学基金项目(81973348);浙江大学实验技术研究项目(SJS202016)。

摘　　要：目的:建立金黄地鼠血浆和组织重组人干扰素α2b定量ELISA检测法,用于药物研发检测。方法:采用双抗夹心法建立金黄地鼠血浆和组织重组人干扰素α2b含量检测方法,确定其检测范围,验证其选择性、精密度与准确度和稀释线性,并对样本稳定性进行研究。结果:建立的方法检测范围为0.50~16.00 ng/ml。选择性结果显示不同个体来源空白金黄地鼠组血浆和组织匀浆无差异,血浆质控样本准确度为78.62%~104.69%,批内精密度为0.69%~18.21%,批间精密度为6.71%~15.81%,总误差均<30%,稀释线性良好。稳定性样本在3种不同保存条件(冰上放置2 h;−80~−75℃放置4 d;反复冻融3循环)下稳定性较好,回收率均在±20%内。结论:成功建立了基于配体结合的人干扰素α2b ELISA检测法,该方法灵敏度高,重复性好,检测效率高,为新药研发提供了可靠的检测方法。Objective:To establish a quantitative ELISA for detection of recombinant human interferon alpha 2b in plasma and tissue of LVG hamsters for drug research.Methods:Double antibody sandwich method was performed to establish detection method of recombinant human IFN alpha 2b in LVG hamster plasma and tissue.Linear range,selectivity,precision,accuracy and dilution linearity of the established detection method were determined.Stability of samples was also detected.Results:Linear range of the established method was 0.50~16.00 ng/ml.Selectivity verification results showed no difference among blank plasma and tissue homogenate samples of LVG hamsters.Accuracy of quality plasma control samples was 78.62%~104.69%,intra assay precision ranged from 0.69%to 18.21%,and inter assay precision ranged from 6.71%to 15.81%.Total method errors were all<30%.Good linear relation was observed in different fold of dilution.Stability samples were stable under three different storage conditions(2 h on ice;−80~−75℃for 4 d;three cycles of freeze-thaw),and recovery was within±20%.Conclusion:Human IFNα2b ELISA method is successfully established based on ligand binding with high sensitivity,good repeatability and high efficiency,providing a reliable detection method for new drug research and development.

关 键 词：重组人干扰素Α2B ELISA 配体结合 方法学验证 

分 类 号：R446.61[医药卫生—诊断学]